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Intratesticular benign peripheral nerve sheath tumour in a ferret (Mustela
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putorius furo) - case report
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Summary
A domestic ferret was submitted for sterilisation due to right testis enlargement.
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Estradiol and cortisol concentrations were within normal physiological ranges, but
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testosterone was below and progesterone above normal. Microscopically, the right testis, with
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the exception of a small part of the epididymis, was replaced with neoplastic tissue. The
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tumour was composed of streams and bundles of closely packed spindle to ovoid cells
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forming whorls around collagen and capillaries, and separated by a collagenous matrix. In
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some areas, cells were loosely arranged and separated by a pale myxomatous matrix. The left
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testis showed atrophy. The majority of neoplastic cells expressed vimentin and S-100 protein
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while expression of collagen IV was moderate and there was no expression of glial fibrillary
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acid protein. On the basis of macroscopical and histopathological findings, and supported by
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immunohistochemical reactivity, the diagnosis of benign peripheral nerve sheath tumour
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(PNST) was made. This is the first report of benign PNST in ferret testis.
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Key words: PNST, ferret, testis, immunohistochemistry, hormone
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Introduction
Peripheral nerve sheath tumours (PNST) are neoplasms arising from Schwann cells,
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perineurial cells, and intraneural fibroblasts. On the basis of morphological features and
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assessment of benignancy versus malignancy, they can be subdivided into schwannoma or
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benign PNST (previously called neurilemoma or neurinoma), neurofibroma or malignant
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PNST (Schulman and others 2009). Due to their uncertain histiogenesis, PNSTs in veterinary
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medicine are divided into benign and malignant variants in the most recent edition of the
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WHO International Histological Classification of Tumours of Domestic Animals (Koestner
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and others 1999). There are few studies of the incidence of neoplasia in ferrets (Brown 1997;
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Zeeland and others 2006). The most common tumours reported are insulinoma, lymphoma
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and adrenocortical neoplasms, followed by skin neoplasias and musculoeskeletal neoplasms
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(Dillberger and Altman 1989). Ovarian stromal tumours are the main neoplasia of the
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reproductive tract in female ferrets, while neoplasms of the male reproductive system are
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unusual (Beach and Greenwood 1993). A few studies have described findings of testicular
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neoplasia in ferrets: interstitial cell adenoma (Meschter 1989), Sertoli cell tumour in
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chryptorchid testis (Powers and others 2007) and mixed interstitial and Sertoli cell tumour
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(Batista-Arteaga and others 2011). In the available literature, there is only a single report of
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PNST in ferret, i.e. neurilemoma on the leg (Brown 1997). We now present the first case of
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benign PNST in a testicle of a ferret and discus in detail the reasons for this diagnosis.
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Case History
A 2.2 kg, male, 6-year old, moderately obese domestic ferret from Zagreb Zoo, was
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submitted to the Surgery, Orthopaedics and Ophthalmology Clinic, Faculty of Veterinary
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Medicine, University of Zagreb in October 2010 for sterilisation due to right testis
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enlargement. During the operation, blood samples were taken for determination of estradiol,
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progesterone, testosterone and cortisol concentrations. Ultrasound evaluation of the abdomen
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was performed prior to the orchidectomy, which was performed through a scrotal incision
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with ligation of the spermatic cord the standard manner.
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The testes were submitted to the Department of Veterinary Pathology for
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histopathological examination. The left testis was round and about 0.8cm in diameter (volume
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of cca. 0.27 cm3), while the right testis had a diameter of about 3.5 cm (volume of cca. 22.43
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cm3). For comparison, the average testicular diameter in sexually mature ferrets is around
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1.2cm (Sisk 1990), with an average volume of 0.94-1.38 cm3 (Schoemaker and others 2008).
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On the basis of these reported values, the left testis (0.8cm, 0.27 cm3) was slightly below the
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normal range, but the right considerably above (3.5cm, 22.43 cm3). The left testis was
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moderately firm and showed no macroscopic changes on the external or cut surface. The right
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testis was firm with a rugged, external surface and was whitish-grey and lobulated on the cut
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surface. The testes were fixed in 10% neutral buffered formalin, embedded in paraffin and
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processed routinely. Sections were stained by haematoxylin and eosin (HE).
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Immunohistochemistry was performed for exact tumour classification. Primary
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antibodies used were specific for the following antigens: vimentin (VIM) (Clone V9,
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Dakocytomation, No. M0725, diluted 1:200), S-100 protein (S-100) (DakoCytomation, No.
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Z0311, diluted 1:400), glial fibrillary acid protein (GFAP) (clone 6F2, DakoCytomation, No.
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M0761, diluted 1:300) and collagen IV (col IV) (clone CIV 22, DakoCytomation, No.
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M0785, diluted 1:25). Antigen retrieval for S-100 was not performed and for VIM was
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performed with microwave pretreatment with proteinase K (DakoCytomation, Code S3004) 5
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min. Antigen retrieval for col IV was performed with ethylene diamine tetraacetic acid
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(EDTA) buffer, pH 9 (DakoCytomation, Code S2367) 3 x 5 min. and for GFAP with citrate
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buffer pH 6 (DakoCytomation, Code S2031) 20 min. in a Dako PT Link module. Endogenous
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peroxidase activity was blocked by 5-min. treatment with hydrogen peroxide (Dako No.
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S2023). Dako REALTM EnVisionTM Detection System, Peroxidase/DAB, Rabbit/Mouse (No.
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K5007) was used to detect the antibodies against VIM, S-100, col IV and GFAP.
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Standard electro-luminiscence serum hormone testing showed an estradiol
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concentration of 5.00 ng/L, progesterone of 18.6 nmol/L, testosterone of 0.087 nmol/L and
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cortisol of 40.8 nmol/L. Ultrasound examination of all abdominal organs, prostate and both
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adrenal glands (left = 7.6 mm length, right = 7.8 mm length) appeared normal (reference
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values reported by Neuwirth and others 1997, are: left = 8.6±1.2 mm; right = 8.9 ±1.6 mm).
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Microscopically, with the exception of a small part of epididymis, the remainder of the
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right testis was replaced with neoplastic tissue (Fig. 1). The tunica albuginea was not
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penetrated by neoplastic cells. Neoplastic tissue was composed of interlacing streams and
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bundles of closely packed spindle to ovoid neoplastic cells and separated by small amounts of
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collagenous matrix. These cells often formed whorls around the collagen and capillaries
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(Antoni A like areas) (Fig. 2). In some areas, cells were more loosely arranged and separated
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by a pale amphophilic, myxomatous matrix (Antoni B like areas) (Fig. 3). Borders of
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neoplastic cells were variably distinct and the cytoplasm was scant and eosinophilic. Nuclei
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varied from round to elongate with finely stippled, marginated or condensed chromatin, with
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one to two poorly distinct nucleoli. Mitotic figures were less than 1/10 HPF. Areas of
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coagulative necrosis and haemorrhage were scattered throughout the tumour. The left testis
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showed marked atrophy. All of the aforementioned microscopic features, except for necrosis
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and haemorrhage, suggested a benign neoplasm. Furthermore, the shape of the cells and their
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arrangement clearly indicated a neoplasm from the group of spindle cell tumours. The two
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different patterns with typical Antoni A and Antoni B like areas with the accompanying
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histological features are characteristic of benign PNST (Bergmann and others 2009; Chijiwa
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and others 2004). Although there were areas of coagulative necrosis and haemorrhage within
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the tumour, there were no other histological criteria of malignancy (infiltrative cell growth
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and aggressive behaviour, presence of atypical mononuclear or multinucleated cells, cellular
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pleomorphism, high mitotic index) to make a diagnosis of malignant PNST (Chijiwa and
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others 2004). Finally, even though histological features of the tumour were clearly suggestive
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of benign PNST, to exclude other spindle cell tumours, IHC stains for VIM, S-100 and col IV
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were applied.
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IHC staining for VIM was strongly positive in cytoplasm (Fig. 4) and S-100 was
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moderately positive in the nuclei and cytoplasm of majority of neoplastic cells (Fig. 5). Col
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IV was moderately extracellularly positive (Fig. 6) whereas staining for GFAB was negative
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in the tumour.
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Discussion
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To our knowledge, this is the second report of a benign PNST in a domestic ferret and first
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report of this tumour in the testis (Brown 1997). The diagnosis of benign PNST was based on
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macroscopic and histopathological findings which were identical to those described in the
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literature (Bergmann and others 2009; Chan and others 2007; Chijiwa and others 2004;
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Koestner and Higgins, 2002; Rothwell and others 1986). Since there is no single specific IHC
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marker that is able to define PNST, a panel of reagents is generally employed. PNSTs are
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mesenchymal spindle cell tumours and are in 100% of cases positive for vimentin (Bergmann
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and others 2009; Chijiwa and others 2004). S-100 protein identifies Schwann cells hence,
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most PNSTs express this molecule (Bergmann and others 2009; Chijiwa and others 2004).
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Since Col IV is a basement membrane component of Schwann cells, PNSTs can also be
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positive to this antibody (Gaitero and others 2008). Finally, GFAB - which is a neuronal
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marker, may be variably expressed (around 67%) in PNSTs (Bergmann and others 2009;
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Chijiwa and others 2004). Spindle cell tumours which can have similar IHC reactivity are
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malignant PNST, rabdomyosarcoma and other tumours of vascular origin (Bergmann and
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others 2009; Chijiwa and others 2004). In our case, typical histological findings excluded
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potential diagnosis of these tumours. Morover, IHC reactivity to VIM, S-100 protein and col
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IV confirmed the diagnosis of benign PNST (Chan and others 2007; Gaitero and others 2008;
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Koestner and Higgins 2002; Schulman and others 2009).
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Estradiol (5.00 ng/L) and cortisol (40.8 nmol/L) levels were within the reference
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ranges of 8±3 ng/L (Carroll and Baum 1989) and 53±42 nmol/L (Rosenthal and Peterson
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1996), respectively. However, the progesterone level (18.6 nmol/L= 5.85ng/mL) was
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approximately twice the normal value (3.1±0.42 ng/mL) (Carlson and Rust 1969) and the
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testosterone level (0.087 nmol/L) was more than 100 times lower than the normal value (9-73
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nmol/L) (Schoemaker and others 2008).
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Since PNSTs are not hormone-secreting neoplasms, this raises some questions
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concerning the altered plasma levels of progesterone and testosterone. One explanation could
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be that progesterone is produced, in male animals, mainly by the adrenal glands and is then
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converted to testosterone in testes (Eckstein and others 1987; Maitra and Abbas 2005).
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Elevated progesterone and decreased testosterone level could be result of replacement of the
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testicular tissue of the right testis with neoplastic tissue and left testis atrophy. Since a very
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small amount of functional testicular tissue remained, a very low conversion rate of
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progesterone to testosterone could be assumed. As a result, testosterone levels would drop and
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progesterone concentration be increased in plasma.
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Considering that surgical castration of male ferrets is a common practice to prevent
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reproduction and to reduce interspecies aggression (Schoemaker and others 2008) it may be
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difficult to define the prevalence of testicular tumours in ferrets (Batista-Arteaga and others
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2011). According to this, our finding of intratesticular benign PNST can contribute in a better
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understanding of the incidence and clinic significance of testicular neoplasm in ferrets.
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Acknowledgements
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This work was supported by Republic of Croatia, Ministry of Science, Education and Sports,
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project no. 053-053-2264-2260.
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Figures:
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Fig. 1. Right testis, ferret. With the exception of a small part of the epididimis, the rest of the
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right testis was replaced with neoplastic tissue (H&E, x10).
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Fig. 2. Right testis, ferret. Antoni A like areas, neoplastic cells form whorls around collagen
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and capillaries (H&E, x10).
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Fig.3. Right testis, ferret. Antoni B like areas, neoplastic cells loosely arranged and separated
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by a pale amphophilic, myxomatous matrix (H&E, x10).
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Fig.4. Right testis, ferret. Neoplastic cells with strong cytoplasmic staining for VIM (IHC,
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x40).
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Fig.5. Right testis, ferret. Neoplastic cells are moderately positive in nuclei and cytoplasm for
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S-100 (IHC, x40).
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Fig.6. Right testis, ferret. A moderate extracellular col IV positive reaction (IHC, x20).